Abstract
The most common numerical chromosome aberration in childhood ALL and NHL is the gain of an extra X chromosome in both male and female patients. We were therefore interested to investigate whether this non-disjunction event affects the active and inactive X chromosomes in a random or non-random fashion. In female cases both the active or inactive X may be duplicated randomly or non-randomly, whereas in male patients only the solitary active homologue can be copied. However, in theory a duplicated active X might subsequently also be subjected to de novo inactivation in both sexes. The inactivation status of acquired X chromosomes is usually evaluated by methylation-specific PCR (MS-PCR), which allows the simultaneous quantification of various differentially methylated polymorphic DNA sequences on the X chromosome, such as those contained in the HUMARA or FMR1 genes. Previous evidence from such analyses suggested that in NHL patients the acquired X chromosomes are and remain always active in male patients, whereas in females both the active and inactive X are duplicated in a random fashion (McDonald et al, Genes, Chromosomes & Cancer 28:246;2000). In childhood ALL this issue has not yet been investigated. However, quantification with MS-PCR has its limitations, especially in cases with low blast cells numbers. To overcome this problem, we have therefore developed a simultaneous dual-color DNA/RNA FISH assay that enables the enumeration of active and inactive X chromosomes on a single cell level. FISH was performed with probes specific for the X centromere and the XIST RNA, which is exclusively expressed from and covers vast parts of the inactive X in human interphase cells. Following the successful evaluation of the assay on methanol/acetic acid-fixed cells that were obtained from 10 healthy individuals and 23 cases with various constitutional X chromosome aneuploidies, we analyzed 54 methanol/acetic acid-fixed samples from hyperdiploid cases of childhood ALL and 29 from NHL. The ALL cases comprised 24 males with two X, 23 females with three X and seven females with four X. The NHL cases consisted of 18 male (9 in the hypo- to hyperdiploid and 9 in the pseudotriploid to pseudotetraploid range) and 11 female patients (7 with three X and 4 with four X chromosomes). In contrast to all constitutional control samples, which as expected contained only one active X, two of the three X in leukemic cell samples from both male and female patients were active. The only exception was a male patient, who most likely was a Klinefelter syndrome with a constitutional XXY. In contrast, all female patients with four X had duplicated both the active and inactive X chromosome. These findings prove that irrespective of the sex of the patient, the active X is exclusively duplicated in cases with three X chromosomes. The consistent gain of both the active and inactive X in female cases with four X, on the other hand, further corroborates previously established evidence that in all instances a single non-discjunction event leads to the maldistribution of chromosomes irrespective of the ploidy range. Moreover, the exclusive presence of duplicated active X chromosomes in hyperdiploid ALL concurs with and explains the results of gene expression profiling studies, which have shown a corresponding over-expression of X-encoded genes.
Identifying (E)-4,8-Dimethyl-1,3,7-Nonatriene Plus Acetic Acid as a New Lure for Male and Female Codling Moth (Lepidoptera: Tortricidae)
